Transplantation arteriosclerosis has continued to limit survival in cardiac allograft recipients. Endothelial cells (ECs) can be primary stimulators and targets of immunologic injury, of which TA may be a chronic form. The long term objective of this work is to define novel molecular interactions between human lymphocytes and allogeneic ECs, and ultimately apply this information to the understanding of immune-mediated vascular injury. Resting T cells react minimally with non-activated endothelium. Natural killer (NK) cells require little antigen-priming and do interact with resting ECs in vitro. The underlying hypothesis is three-fold: (1) immune-mediated vascular injury is a consequence of cytotoxic lymphocyte stimulation/activation by ECs, and EC apoptotic death; (2) ECs are antigen-presenting cells (APC) and their ability to drive lymphocyte activation is enhanced by engagement of beta2 integrins, most notably LFA-1 with EC ligands ICAM-1 and -2; and (3) despite APC activity, EC are intrinsically deficient in their ability to present class I HLA-associated peptides, providing the """"""""substrate"""""""" for allorecognition of the endothelium by and sensitivity to NK lymphocytes. Cytotoxic lymphocyte-EC adhesion, mutual cellular activation and/or EC lysis are highly interdependent phenomena which may constitute the cellular triggers for vascular injury. Based on the investigators' prior observations of exquisite sensitivity to NK lymphocyte-mediated lysis via apoptotic EC death, the EC protection conferred by IFNg in this apoptotic response, and the important role of LFA-1 in this phenomenon, specific proposals now include (1) definition of LFA-1 mediated signals that confer stabilization to lymphocyte activation mRNAs, including (a) dependence on the cytoskeleton and small GTP-binding proteins, and (b) modulation of RNA binding proteins; (2) identification of class I-associated peptides on the EC membrane and differences between these peptides in resting, NK-sensitive vs IFNg-activated, NK-resistant EC in order to define a class of protective peptides; (3) determining the mechanisms of NK cell-induced EC apoptosis, focusing on the role of perforin and granzyme B activation of ICE proteases, and addressing modulatory functions of IFNg in these responses; and (4) characterization of a novel IFNg-activated gene, UBDF, which the investigators cloned and which encodes a protein with ubiquitin and DAF homology domains, and assess its protective potential against lymphocyte-mediated EC apoptosis.
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